The long term objective of this proposal is to elucidate the function and regulation of telomerase in normal and cancer cells. Telomerase is the enzyme that replicates the DNA ends, or telomeres, of chromosomes. Normally this enzyme is repressed in most human cells, resulting in a loss of terminal DNA every cell division. This erosion of telomeres is ultimately lethal, and thus limits the lifespan of such cells. However, during the process of tumourigenesis cancer cells acquire the ability to proliferate well beyond the replicative capacity of normal cells. We and others have now shown that cancer cells appear to surmount the proliferative barrier imposed by telomere shortening through the upregulation of telomerase via the illegitimate activation of the gene hTERT, which encodes the catalytic subunit of telomerase. However, very little is know how this telomerase protein elongates telomeres, and what other proteins participate in this reaction. Additionally, mounting evidence suggests that a variety of cellular pathways may converge upon telomerase, but again how such pathways may regulate telomerase activity is uncertain. To address these issues two aims are proposed that capitalize on our previous biochemical and genetic analysis of the telomerase catalytic subunit in humans and yeast: 1. Map and characterize the functional domains of the catalytic subunit of telomerase. Identify and characterized proteins that interact with these domains. 2. Identify and characterize the human homologues of yeast proteins known to be important for telomerase function. The accomplishment of the above aims will further our understanding of how telomerase functions, of how this function is controlled and on what signals regulate this enzyme activity. Such information would be invaluable in understanding the role of telomerase in cellular immortalization which in turn could lay the foundation for the development of agents to inhibit telomerase function, which ultimately may be of therapeutic value in the treatment of human cancers.